NOX2-mediated reactive oxygen species contribute to angiotensin II-induced cardiac hypertrophy by promoting cardiomyocyte autophagy via Atg4B S-glutathionylation modification

The NADPH Oxidase (NOX) 2-dependent Reactive Oxygen Species (ROS) mediate cell Autophagy to participate in a variety of cardiovascular diseases. However, whether NOX2-mediated ROS facilitates angiotensin II-induced cardiac hypertrophy by promoting cardiomyocyte Autophagy is still unknown. Mouse cardiomyocytes were stimulated with angiotensin II for different time in the presence and absence of apocynin or 3-methyladenine. Angiotensin II-induced NOX2-knockdown cardiomyocytes and NOX2-deficient mice were used to explore the function of NOX2 in angiotensin II-induced cardiomyocyte hypertrophy, oxidative stress (OxS), and Autophagy. The abundance of several proteins was detected by western blot. Wheat germ agglutinin and Masson's staining determined cardiomyocyte size and cardiac fibrosis. OxS was determined by analyzing ROS production, malondialdehyde levels, and superoxide dismutase activity. Co-immunoprecipitation was performed to explore the S-glutathionylation of autophagy-related gene 4B (Atg4B). Apocynin and 3-methyladenine treatment decreased angiotensin II-mediated elevation in atrial natriuretic peptide and LC3 II/LC3 I protein levels but not NOX2. NOX2 silencing impaired angiotensin II-induced cell hypertrophy, ROS production, and Autophagy in cardiomyocytes. Consistently, NOX2-deficient mice exhibited improvements in cardiac dysregulation, hypertrophy, and fibrosis, coupled with reduced OxS and Autophagy. NOX2 silencing decreased angiotensin II-induced upregulation of Atg4B. Moreover, Atg4B knockdown improved angiotensin II-induced cardiomyocyte hypertrophy and Autophagy, whereas NOX2 overexpression did not work in Atg4B knockdown-cardiomyocytes. Mechanistically, angiotensin II-induced NOX2 facilitates the S-glutathionylation of Atg4B. NOX2-dependent ROS facilitates angiotensin II-induced cardiac hypertrophy via Atg4B S-glutathionylation modification, indicating that targeting the S-glutathionylation of Atg4B may be a potential adjunct therapeutic strategy for cardiac hypertrophy, complementary to existing standard treatments.

Autophagy; Cardiac hypertrophy; Mice; NADPH oxidase; ROS.